Blood, spinal fluid, cell cultures and urine are examples of biological fluids which are microscopically examined to determine the presence or concentration of suspended particulate matter such as cells. For many years microscopic examination of liquid specimens had been accomplished by placing a drop of the specimen liquid on a flat transparent microscope slide and then placing a thin flat transparent cover slip over the specimen. More recently preassembled plastic slides with at least one examination chamber formed between a base plate and cover plate have been used. Liquid specimens are introduced into the examination chambers by placing a drop of the liquid specimen adjacent to a chamber and having capillation draw the specimen into the chamber. Preassembled plastic slides of this type are taught in U.S. Pat. No. 4,637,693 issued Jan. 20, 1987.
In order to accurately determine the concentration of suspended particles or cells in a specimen, several parameters must be either measured or maintained at constant values. These parameters are the interior dimensions of the volume of the examination chamber where suspended particles or cells are being counted. It is taught in U.S. Pat. No. 4,637,693 how to make plastic slides with the roofs and floors of examination chambers fixed in parallel relationship to each other at a predetermined distance. Since the roofs and floors are parallel and at a fixed distance the only remaining dimension required in determining the volume of specimen liquid being examined is the lateral dimension of the field of view of the examining instrument, i.e., microscope. This lateral dimension varies depending on magnification and the optics used in the microscope. Therefore, standardization when using multiple microscopes requires either using microscopes having the same fields of view or calculating the differences in fields of view and factoring the differences into the counts of suspended particulate matter.
An alternative to working with the lateral extent of the field of view while still utilizing the same preassembled plastic slide would be to incorporate a grid system on the floor of the examination chamber. The preassembled slides are made of transparent optical quality plastic and therefore the grid must either be machined or scribed onto the floor plate of the slide after molding or must be a pattern incorporated in the mold used to make the base portion of the slide. The most efficient process would be to have the grid pattern incorporated in the mold.
Many applications require magnification of the order of 400 times. At this magnification the diameter of the fields of view vary from about 0.33 millimeters (mm) to 0.50 mm. So useful grid systems must be set up with defining lines separated by less than half a mm and accordingly very narrow lines such as 0.025 mm wide should be used. Otherwise, portions of the grid perimeter will extend beyond the field of view requiring adjustments to the microscope thus effectively defeating the purpose of the grid.
Previously, grids have been directly etched on glass slide bases for hemacytometers with lines of 0.013 mm width. However, up to the time of the present invention, trying to get such fine line widths on metal mold parts used to make preassembled plastic slides have not been successful. For example, line widths could only be reduced to 0.076 to 0.13 mm using milling machine and tool bit techniques.